Determination of amphetamine in dog plasma by gas chromatography with mass selective detection

Abstract

This paper describes the validation of an analytical method for the determination of amphetamine in beagle dog plasma by gas chromatography coupled to mass spectrometry (GC-MS). d-Amphetamine-d(6) was used as the internal standard. The method consisted of a rapid single-step liquid-liquid extraction and derivatization of amphetamine with 2,2,2-trichloroethyl chloroformate, followed by sensitive GC-MS detection. This is the first report utilizing the combination of trichloroethyl chloroformate as a derivatization reagent and a deuterated amphetamine analog as an IS for the quantification of amphetamine in plasma. The method was validated in terms of specificity, curve fit, precision, accuracy, recovery and stability, and was acceptable according to FDA draft guidelines for validation of bioanalytical methods. The limit of detection was 0.65 ng/mL. The calibration range was 5-150 ng/mL. The validated method was successfully employed for the quantitation of amphetamine in dog plasma samples for pharmacokinetic profiling.

Development and validation of a liquid chromatography/tandem mass spectrometry assay for the simultaneous determination of d-amphetamine and diphenhydramine in beagle dog plasma and its application to a pharmacokinetic study

“…Studies on the pharmacokinetics of diphenhydramine have been reported in rabbits [9], sheep [20], human beings [19], camels and horses [33], except beagle dogs. There is only one literature on the pharmacokinetics of amphetamine in beagle dogs, reported by March et al [11], and the pharmacokinetic parameters after an oral administration of 10 mg of 'immediate- 11.6 ± 3.9 6.5 ± 0.8 AUC 0− (g h/mL) 3.8 ± 1.6 9.2 ± 3.1 AUC 0−∞ (g h/mL) 4.1 ± 2.1 9.5 ± 3.2 release' amphetamine dose are as follows: C max = 86.6 ng/mL, T max = 1 h, AUC 0− = 0.56 g h/mL, AUC 0−∞ = 0.67 g h/mL. In our study, the concentration of d-amphetamine increasd gradually up to 450.1 ng/mL at 2.3 h after an oral administration of 4 QAAMCs, corresponding to 20 mg d-amphetamine, AUC 0− and AUC 0−∞ are 3.8 and 4.1 g h/mL, respectively.…”

“…Therefore, to further understand and reveal the pharmacokinetic profile and mechanism of the QAAMC, sensitive and specific analytical methods for the simultaneous determination of d-amphetamine and diphenhydramine were urgently needed. Some procedures have been described for the determination of either d-amphetamine or diphenhydramine in biological fluids, such as gas chromatography-mass spectrometry [9][10][11], capillary electrophoresis with UV-absorbance detection [12][13][14][15], HPLC with UV-absorbance detection [16][17][18], however, these methods do not meet modern drug development needs with respect to an efficient extraction procedure, shorter run time and high sensitivity. Liquid chromatography coupled to tandem mass spectrometry has become a method of the choice for the determination of small molecules in biological matrices, including d-amphetamine and diphenhydramine [19][20][21][22], because of its superior LLOQ, sensitivity, and improved selectivity.…”

Gas chromatography/mass spectrometry determination of amphetamine‐related drugs and ephedrines in plasma, urine and hair samples after derivatization with 2,2,2‐trichloroethyl chloroformate

“…With the sole exception of the already described AM and MA derivatives,33, 34 this paper reports, to the best of our knowledge, the first application of 2,2,2‐trichloroethyl chloroformate derivatization of ARDs and ephedrines. Figure 1 shows the general derivatization reaction of the analytes and the common EI mass fragmentation pathways of their 2,2,2‐trichloroethylcarbamate derivatives.…”

“…2,2,2‐Trichloroethyl chloroformate has already been used to derivatize several acid and alkaline compounds25–32 and, although also employed for determination of AM and MA in biological samples,33, 34 to the best of our knowledge no reports of its extensive application in ARD bioanalysis are available to date.…”

Surface‐activated chemical ionization ion trap mass spectrometry in the analysis of amphetamines in diluted urine samples

“…Amphetamines can be usually detected in urine, plasma samples, nails or hair samples 5–7. Several different analytical techniques are routinely employed for the analysis of these compounds, based on the immunochemical approach (radioimmunoassay (RIA), enzyme immunoassay (EIA), and fluorescence immunoassay polarization (FPIA)) 5.…”